新疆维吾尔族妇女子宫颈癌活检组织中人类乳头状…

目的：研究我国宫颈癌高发区新疆维吾尔族妇女宫颈癌与人类乳头状瘤病毒（ＨＰＶ）感染的关系。方法：对６５便新疆维吾尔族妇女的宫颈癌活检组织标本，应用原位杂交法检测ＨＰＶ６／１１、１６／１８和３１／３３／３５ＤＮＡ；Ｌ１共有序列引物聚合酶链反应（ＰＣＲ）及Ｅ６特异型引物ＰＣＲ检测其中的５８例ＨＰＶ６、１６和１８ＤＮＡ。结果：６５例宫颈癌患者中，原位杂交法检测ＨＰＶＤＮＡ有２８例阳性（４３．１％）；Ｌ１Ｐ     

子宫颈癌p53抑癌基因突变与病毒感染的研究

采用聚合酶链反应－单链构象多态性分析（ＰＣＲ－ＳＳＣＰ）、多重引物ＰＣＲ和巢式引物ＰＣＲ，对同一宫颈癌组织中ｐ５３基因（第６～８外显子）突变以及人乳头状瘤病毒（ＨＰＶ）和人巨细胞病毒（ＨＣＭＶ）感染进行相关性研究。并与正常宫颈组织进行对照。结果：３８例宫颈癌组中，２例有ｐ５３第７外显子突变。其中１例伴有ＨＰＶ１６感染，２例伴有ＨＣＭＶ感染。宫颈癌组ＨＰＶＩ６、１８ＤＮＡ的阳性率为６３．２％（２４／３８），ＨＣＭＶＤＮＡ为８４．２％（３２／３８）。２１例对照组中，ＨＰＶ１６、１８ＤＮＡ和ＨＣＭＶＤＮＡ的阳性率分别为４．８％和３８．１％（Ｐ＜０．００５）。ＨＰＶ１６、１８阳性的子宫颈癌中，８７．５％伴有ＨＣＭＶ感染。对照组中，无一例同时检测出ＨＰＶ１６、１８和ＨＣＭＶ者。提示：宫颈癌组织中，ｐ５３基因突变并不常见，其突变与ＨＰＶ１６、１８感染无显著关系。宫颈癌与ＨＰＶ１６、１８关系密切，ＨＣＭＶ可能与ＨＰＶ协同作用，导致宫颈癌的发生。     

人乳头状瘤病毒感染与子宫颈癌发病关系的探讨

应用多重引物人乳头状瘤病毒（ＨＰＶ）６Ｂ／１１、１６、１８　型聚合酶链反应（ＰＣＲ）技术检测９９例不同宫颈病变宫颈组织中人乳头状瘤病毒ＤＮＡ（ＨＰＶＤＮＡ）。其中宫颈湿疣２０例，宫颈上皮内瘤变（ＣＩＮ）１～２级（ＣＩＮ＿（１～２））１８例，ＣＩＮ３级（ＣＩＮ＿３）２０例，宫颈癌２３例和正常对照１８例。结果：上述宫颈组织中ＨＰＶＤＮＡ总检出率分别为８５．０％，８３．３％，８０．０％，８７．０％和２７．８％。宫颈各病变组中ＨＰＶＤＮＡ的检出率均显著高于对照组（Ｐ＜０．０１）。宫颈湿疣及ＣＩＮ＿（１～２）中ＨＰＶ６Ｂ／１１型检出率分别为８５．０％和７２．３％，ＣＩＮ＿３和宫颈癌中ＨＰＶ６和（或）１８型阳性率分别为５０．０％和７３．９％，两者ＨＰＶ型别分布差异有显著意义（Ｐ＜０．０１）。３例腺癌中ＨＰＶ１８型阳性２例，１６型阳性１例。低分化宫颈癌中均为ＨＰＶ１６和（或）１８型感染。提示：宫颈癌及其癌前病变的发生与ＨＰＶ感染高度相关。宫颈湿疣和ＣＩＮ＿（１～２）常与ＨＰＶ６Ｂ／１１型感染有关；ＣＩＮ＿３和宫颈癌的发生则与ＨＰＶ１６、１８型关系密切。     

子宫颈鳞癌组织中人乳头状瘤病毒E7蛋白与Rb基因产物的关系

目的：检测子宫颈鳞癌组织中是否存在人乳头状瘤病毒（ＨＰＶ）Ｅ７蛋白与Ｒｂ基因产物（ｐＲｂ）的结合物（ＨＰＶＥ７－ｐＲｂ）。方法：运用聚合酶链反应（ＰＣＲ）技术，对４０例宫颈鳞癌组织中ＨＰＶ６／１１、１６、１８、３３型ＤＮＡ进行检测，并用捕获酶联免疫吸附试验，探查ＨＰＶ感染的标本中有否ＨＰＶＥ７－ｐＲｂ存在。结果：４０例中，检出ＨＰＶＤＮＡ１８例（４５．０％）。１８例中９例存在ＨＰＶＥ７－ｐＲｂ，其中１例为ＨＰＶ１８，８例为ＨＰＶ１６。２例ＨＰＶ６／１１未检出ＨＰＶＥ７－ｐＲｂ。临床Ⅰ期患者ＨＰＶＥ７－ｐＲｂ的检出率高于Ⅱ～Ⅳ期患者（Ｐ＜０．０５）。ＨＰＶＥ７与ｐＲｂ的结合，与宫颈癌病理分级无关（Ｐ＞０．０５）。结论：在宫颈鳞癌组织中存在ＨＰＶＥ７－ｐＲｂ；ＨＰＶＥ７与ｐＲｂ的结合过程发生在宫颈癌变的较早阶段。     

宫颈癌中HPV16 E6蛋白诱导内质网应激-自噬反应的研究

目的：研究宫颈癌Ｃ３３Ａ细胞中ＨＰＶ１６ Ｅ６蛋白过表达对内质网应激－自噬反应的影响。方法：构建ＨＰＶ１６ Ｅ６的真核表达载体ｐｃＤＮＡ３．１（－）－ＨＰＶ１６ Ｅ６,转染Ｃ３３Ａ细胞［ｐｃＤＮＡ３．１（－）－ＨＰＶ１６ Ｅ６组］,同时设空载体转染组［ｐｃＤＮＡ３．１（－）组］和空白对照组（Ｃ３３Ａ组）,荧光定量聚合酶链反应（ＰＣＲ）和蛋白质印迹法（Wｅｓｔｅｒｎ ｂｌｏｔ）检测其Ｃ３３Ａ细胞中ＨＰＶ１６ Ｅ６ ｍＲＮＡ及蛋白表达的变化,及其对自噬蛋白Ｂｅｃｌｉｎ １、ＬＣ３Ⅱ、内质网应激标记蛋白葡萄糖调节蛋白78（ＧＲＰ７８）表达的影响,四甲基偶氮唑盐（ＭＴＴ）法检测细胞生长的变化;流式细胞仪检测细胞自噬的变化。结果：成功构建了ＨＰＶ１６ Ｅ６的真核表达载体ｐｃＤＮＡ３．１（－）－ＨＰＶ１６ Ｅ６,其可以使Ｃ３３Ａ细胞中ＨＰＶ１６ Ｅ６ ｍＲＮＡ及蛋白表达增加;ＨＰＶ１６ Ｅ６在Ｃ３３Ａ中过表达可促进肿瘤细胞的生长,ｐｃＤＮＡ３．１（－）－ＨＰＶ１６ Ｅ６转染后Ｃ３３Ａ的自噬率较ｐｃＤＮＡ３．１（－）组和Ｃ３３Ａ组增高,差异有统计学意义（Ｐ＜０．０５）;ＨＰＶ１６ Ｅ６过表达使Ｂｅｃｌｉｎ １、ＬＣ３Ⅱ和ＧＲＰ７８蛋白表达增加,阻断内质网应激通路后ＧＲＰ７８表达减少,细胞自噬率降低。结论：内质网应激－自噬反应在ＨＰＶ１６感染导致宫颈癌的过程中具有重要作用,这为宫颈癌的基因治疗提供了新靶点。     

应用聚合酶链反应技术测定正常子宫颈组织人乳头瘤病毒DNA的研究

应用兼并复合人乳头瘤病毒一聚合酶链反应（ＨＰＶ－ＰＣＲ）技术对１４８例外观正常的子宫颈分泌物拭子标本进行人乳头瘤病毒（ＨＰＶ）ＤＮＡ的测定。结果：正常子宫颈ＨＰＶＤＮＡ检出率为３１７６％（４７／１４８）。其中ＨＰＶ－６，１１型为６．７６％（１０／１４８）；ＨＰＶ－１６，１８型为３．３８％（５／１４８）；ＨＰＶ其它型为２１．６２％（３２／１４８）。外观正常的子宫颈，ＨＰＶ感染的主要易感年龄为２２～３４岁的青年妇女。本研究对女性生殖道ＨＰＶ感染的传播途径进行了探讨。     

用光敏生物素标记的HPV6，11，16，18四型探针检测宫颈癌组织中的HPV核酸序列

本文运用分子生物学方法，制备了光敏生物素标记的人乳头瘤病毒（ＨＰＶ）６，１１，１６，１８四型探针。用原位杂交法检测了经临床诊断及病理证实为宫颈癌组织申的ＨＰＶＤＮＡ相关序列。在３３例宫颈癌组织中，有６例呈ＨＰＶ６阳性，阳性率为１８．１％；７例呈ＨＰＶ１１阳性，阳性率为２１．２％；２２例呈ＨＰＶ１６阳性，阳性年为６６．７％，１８例呈ＨＰＶ１８阳性，阳性率为５４．５％；１０例正常宫颈组织呈阴性。提示黑龙江地区宫颈癌的发生与ＨＰＶ有关，尤其是与ＨＰＶ１６，１８两型感染关系密切．特别是ＨＰＶ１８型感染率在本组中占有很高的比例。     

女性下生殖道尖锐湿疣的诊断

采用分子生物学多聚酶链反应（ＰＣＲ）方法，对女性下生殖道疣状赘生物６１６例妇女（团块型３０７例，丘疹型３０９例），进行人乳头瘤状病毒ＨＰＶ６，１１，１６，１８，３３，３５ＤＮＡ等４８个亚型检测，并与以免疫组织化学ＡＢＣ方法检测ＨＰＶ－Ａｇ（衣壳抗原）及电镜、光镜、阴道镜等方法进行比较。结果：ＰＣＲ方法的灵敏度和特异性最强，团块型阳性率９７．９０％，丘疹型１．１０％，两型共存者均团块型阳性而丘疹型阴     

尖锐湿疣及子宫颈癌组织人乳头状瘤病毒DNA的比较研究

目的：探讨不同种类人乳头状瘤病毒（ＨＰＶ）感染与病变性质的关系。方法：采用聚合酶链反应和Ｓｏｕｔｈｅｒｎ杂交方法，检测尖锐湿疣８９例（尖锐湿疣组）、宫颈癌７６例（宫颈癌组）和正常生殖道１９８例（对照组）组织标本中ＨＰＶ－ＤＮＡ，并对ＨＰＶ感染的主要类型进行分析。结果：尖锐湿疣组中，ＨＰＶ－ＤＮＡ检出率为９８．９％，ＨＰＶ６和ＨＰＶ１１占其中的８６．４％；宫颈癌组中，ＨＰＶ－ＤＮＡ检出率为９４．７％，ＨＰＶ１６和ＨＰＶ１８为主要类型，占８７．５％；对照组中ＨＰＶ隐匿感染率为３７．４％，ＨＰＶ６、１１、１６、１８仅占２１．６％。尖锐湿疣组织中ＨＰＶ１１－ＤＮＡ主要以游离形式存在，而宫颈癌组织中ＨＰＶ１６－ＤＮＡ绝大多数整合到宿主细胞中，并有某种程度的变异。结论：ＨＰＶ的不同类型与病变的组织学损害所表现的特殊形态及性质改变有关。ＨＰＶ－ＤＮＡ与宿主细胞ＤＮＡ的关系及其变异情况，可确定疾病的性质。     

人乳头状瘤病毒感染对孕妇和绒毛滋养细胞的影响

目的 探讨人乳头瘤病毒（ＨＰＶ）感染对孕妇和绒毛滋养细胞铁影响。方法 观察６２例早、中期孕妇尖锐湿疣的病理形态特点,用ＨＰＶ６／１１－ＤＮＡ探针原位杂交法检测其绒毛、胎盘组织及其中１０例外阴、阴道、宫颈尖锐湿疣组织,并用ＨＰＶ６／１１聚合酶链反应（ＰＣＲ）方法验证１０例绒毛及胎盘组织。结果 孕期尖锐湿疣病变发展快,多部位发生比例高,大体形态、组织形态典型者多。ＨＰＶ６／１１－ＤＮＡ原位杂交和ＰＣＲ     

新疆南部维吾尔族妇女宫颈癌及癌前病变HPV16 E6/E7基因突变序列分析

目的:探讨新疆南部地区维吾尔族妇女HPV16E6/E7基因突变与宫颈病变及宫颈癌发生的关系。方法:提取上述地区40例宫颈癌及80例CINⅠ～Ⅲ组织的HPV16DNA,用PCR扩增其E6/E7基因并对产物测序,然后以德国标准株HPV16DNA为原型进行对比,分析其突变情况。结果:HPV16E6突变中L83V占80.6%(25/31),D25E占16.1%(5/31),E7突变中R77C占33.3%(5/15),N29S占26.7%(4/15)。结论:新疆宫颈病变及其癌变组织中HPV16E6/E7以L83V,D25E,R77C,N29S为主要突变株,欧洲型突变为主,与亚洲型突变同时出现是该地区的主要突变特点。     

HPV16/18 prevalence in cervical lesions/cancers and p53 genotypes in cervical cancer patients from India

OBJECTIVES: The HPV16/18 code for an oncoprotein-E6, which binds to p53 tumor suppressor protein and degrades the protein via ubiquitination. A common polymorphism of p53 in exon 4 codon 72, resulting in either proline (Pro) or arginine (Arg), affects HPV16/18 E6-mediated degradation of p53 protein in vivo. Hence, in the current study we investigated the prevalence of HPV16/18 in cervical lesions and the distribution of p53 genotypes in cervical cancers and normal healthy women. METHODS: DNA from 337 Indian women with invasive cervical cancers, 164 women with clinically normal cervix, 64 women with low-grade squamous intraepithelial lesions (LSIL), and 5 women with high-grade squamous intraepithelial lesions (HSIL) was examined for the presence of HPV16/18 using consensus primers in a polymerase chain reaction (PCR), and the specific HPV type was identified by Southern hybridization of the PCR product using HPV16/18 type-specific nucleotide sequences as probes. Further, 134 women with cervical cancers and 131 healthy women were used to determine the frequency of p53 genotypes, Pro/Pro, Arg/Arg, and Pro/Arg, using peripheral blood cell DNA to indicate the constitutional genotypes and allele-specific primers, in a PCR-based assay. RESULTS: We observed a prevalence of HPV16/18 in 77% (258/337) of cervical cancer patients, 38% (24/64) of LSILs, 4 of 5 HSILs, and 15.2% (25/164) of normal healthy women. The frequency of distribution of the three genotypes of p53 codon 72 in a subgroup of the HPV16/18-positive cervical cancer patients was Pro/Pro 0.18 and Arg/Arg 0.26, with the heterozygous Pro/Arg 0.56, differing significantly from the genotype frequency in the normal healthy women (chi(2) = 6.928, df = 2, P < 0.05). CONCLUSIONS: A high prevalence of HPV16/18 was observed in the cervical cancers. The prevalence in LSILs confirms HPV16/18 infection as an early event and further indicates a role in progression of lesions. The p53 genotype distribution indicated that women homozygous for Arg genotype were at a 2.4-fold higher risk for developing HPV16/18-associated cervical carcinomas, compared to those showing heterozygous Pro/Arg genotype (odds ratio 2.4, 95% confidence interval 1.89 to 3.04).     

A simple method for the detection and genotyping of high-risk human papillomavirus using seminested polymerase chain reaction and reverse hybridization

BACKGROUND: To develop a simple and cost-effective method for the detection and genotyping of high-risk human papillomaviruses (HPV) using seminested polymerase chain reaction (PCR) and reverse hybridization. METHODS: Cervical swabs for HPV testing were collected from 127 women with normal cervical cytology and 57 patients with cervical lesions of various degrees. After DNA isolation, PCR amplification was first carried out using MY11 and MY09/HMB01 primers, then labeled by seminested PCR using the first PCR products and MY11/bioGP6+ primers. One fifth of the second PCR products were resolved by gel electrophoresis. Genotyping for high-risk HPV was done separately, using the remaining products, by a high-risk HPV chip, which contained 13 type-specific oligonucleotides on a nylon membrane. The final result was detected by colorimetric change on the chip under direct visualization. RESULTS: High-risk HPV DNA was detected in 19 (15%) of 127 women with normal cervical smear cytology, in 26 (89.7%) of 29 patients with cervical intraepithelial neoplasia (CIN), and in 27 (96.4%) of 28 patients with invasive cervical carcinoma. Multiple high-risk HPV infections were detected in five cases. HPV type 16 was the most frequent type of infection, comprising 34.5% and 53.6% of the patients with CIN and invasive carcinoma, respectively. The samples without a visible 190-bp band on electrophoresis exclusively showed negative hybridization results. This method could detect one to two copies of the HPV-16 genome derived from one SiHa cell. The overall sensitivity of HPV detection was 25 to 50 copies of HPV genome for each specimen. Thirteen high-risk types and twenty-four different types of HPV DNA showed specific hybridization without any cross-reaction. CONCLUSIONS: Our results demonstrated the feasibility and optimistic prospects for this simple and cheap method of high-risk HPV genotyping. This technology can be easily set up in a routine molecular laboratory and would probably be of great value in cervical cancer prevention programs.     

Detection of HPV DNA in the uterine cervical lesions by polymerase chain reaction and in situ hybridization]

Human papillomavirus (HPV) is found in close association with carcinogenesis of the uterine cervix. We applied a new in vitro gene amplification technology, the polymerase chain reaction (PCR) to detect HPV 16 and 18 in cervical exfoliated cells. HPV infections were detected in 5 (16%) of 31 women with no pathological lesions of the uterine cervix (normal), 16 (24%) of 67 with cervical intraepithelial neoplasia (CIN) and 6 (38%) of 16 with invasive cervical cancer. Moreover, 10% formalin-fixed and paraffin-embedded tissue sections were prepared from the uterine cervix of these 27 women with PCR-proven HPV infection and were examined for the histological localization of HPV-DNA by in situ hybridization with biotin-labeled DNA probes of HPV types 6/11, 16/18 and 31/33/35. HPV-DNA type 16/18 was detected in 3 of 5 normal women, 2 of 4 CINs I, 2 of 3 CINs II, 6 of 9 CINs III and 6 of 6 invasive cervical cancers. HPV-DNA type 6/11 was detected in 6 of 6 condylomas. Viral DNA sequence was detected in the superficial cells of CIN I and II, and it was distributed through entire thickness layer of undifferentiated cells derived from CIN III and squamous cell carcinoma. In addition, the staining intensity became weak as the lesion progressed. These differences between lesions might be due to the difference in the viral form in the nuclei, ie whether an episomal or integrated form. Thus, an in situ hybridization technique with a biotin-labeled DNA probe as well as the PCR method is useful for the detection of HPV in clinical samples.     

Impact of human papillomavirus coinfections on the risk of high-grade squamous intraepithelial lesion and cervical cancer

Objective

The molecular and epidemiologic effect of human papillomavirus (HPV) coinfections in the risk of developing cervical cancer is yet unclear. The aim of this study was to determine the frequency HPV coinfections at different stages of cervical lesions in the development of cervical cancer and the impact of HPV specific type interactions on high-grade squamous intraepithelial lesions (HSIL) and invasive cervical cancer (ICC) risk.

Methods

HPV testing was performed in 931 cervical samples diagnosed as: negative for intraepithelial lesion or malignancy (NILM); low-grade squamous intraepithelial lesion (LSIL); HSIL; and ICC. For HPV detection and typing two sets of primers from the L1 region were used in the polymerase chain reaction method (PCR) (MY09/MY11/HMB01 and L1C1/L1C2.1/L1C2.2) and HPV type was determined by PCR product sequence. To look for multiple HPV infections, the E6 nested multiplex PCR method was performed in all DNA samples. Odds ratios were calculated as indexes of the strength of the association between the sample category (LSIL/NILM or ICC/HSIL) and the presence of a given viral combination.

Results

In HPV positive samples, coinfections are as common in ICC/HSIL as in LSIL/NILM (47.12% and 40.17%, respectively). There is an increased risk to ICC/HSIL when multiple high-risk HPV types are present. The coinfection of HPV68 with HPV16 increases the risk of ICC/HSIL (OR = 14.54, P = 0.012, after multivariate adjustment), related to the presence of HPV16 or HPV68 alone.

Conclusions

These results sustain that specific HPV coinfections confer an increased risk to develop ICC/HSIL.     

Development and evaluation of a highly sensitive human papillomavirus genotyping DNA chip

OBJECTIVES: Test of human papillomavirus (HPV) is a useful adjunctive tool of Pap smear to screen cervical cancer. We have developed a novel HPV genotyping DNA chip arrayed by multiple oligonucleotide probes of both L1 and E6/E7 gene sequence of 42 types of anogenital HPV. METHODS: Consensus PCR products of L1 and E6/E7 gene sequences of HPV are hybridized to arrayed probes on the HPV chip and HPV genotypes are identified by fluorescence scanner. We have comparatively analyzed the value of HPV DNA chip and DNA sequencing in 100 cervical cancer tissues. RESULTS: Overall, 98 cervical cancer tissues were found to harbor DNA sequences of high-risk type HPVs, of which 88 (89.8%) were detected by PCR-sequencing of L1 alone, 98 (100%) by PCR-sequencing of both L1 and E6/E7, and 98 (100%) by HPV DNA chip, respectively. All of the genotypes of HPV detected on sequencing analysis were also found on DNA chip analysis. HPV DNA chip was superior to direct DNA sequencing in detection of mixed infection. CONCLUSIONS: These results suggest that HPV DNA chip analysis in the present study is highly accurate for detection and genotyping of HPV and may have potential value as a robust, high-throughput screening test of uterine cervix cancer.     

Detection of human papillomavirus DNA in malignant lesions from Chinese women with carcinomas of the upper genital tract

OBJECTIVE: The aim of this study was to determine the prevalence of high-risk oncogenic human papillomaviruses (HPVs) in malignant lesions from Hong Kong Chinese women with carcinomas of the upper genital tract. METHODS: The presence of high-risk HPVs in 55 cases of endometrial adenocarcinomas and 60 cases of primary epithelial ovarian cancers was detected by polymerase chain reaction (PCR) using consensus primers complementary to late 1 (L1) gene of the genital HPVs. Amplified PCR products were verified and typed by Southern blot analysis using (32)P-labeled DNA probes prepared from cloned HPV-16 and -18 plasmids. To confirm the presence of high-risk HPV types in the tumor tissues, PCR amplification using HPV type 16- and 18-specific primers for part of the E6 gene were also carried out. RESULTS: While HPV-18 was not detected, HPV-16 DNA sequences were identified in 5 (9.1%) of the 55 studied endometrial carcinoma samples. Of the 5 HPV-16-positive cases, there were 4 stage I, and 1 stage II endometrial cancer. In addition, 6 (10%) of the 60 epithelial ovarian carcinomas were positive for high-risk HPVs, which included 5 cases with HPV-16 and 1 case with HPV-18. Clinical staging revealed that 5 of the 6 HPV-positive cases were stage I and the remaining case was stage III ovarian cancer. Histology of the 6 HPV-positive cases showed that there were 1 case of clear-cell adenocarcinoma, 1 case of mucinous cystadenocarcinoma, and 4 cases of mucinous tumor of borderline malignancy. No other HPV types were detected. CONCLUSION: High-risk HPV was detected in approximately 10% of the tumor samples from women with upper genital tract carcinomas. As compared to the high positive rate of HPV infections in cervical cancer, it appears that HPV infection plays a relatively minor role in the pathogenesis of endometrial and ovarian carcinomas.     

Prognostic significance of polymerase chain reaction detected human papillomavirus of tumors and lymph nodes in surgically treated stage IB cervical cancer

This study describes the prognostic role of polymerase chain reaction detected human papillomavirus (HPV) in Stage IB cervical cancer patients treated with radical hysterectomy and pelvic and paraaortic node dissection. All tumors were confined to the cervix and all margins and nodes were disease free. Twenty-one patients were analyzed: 6 patients recurred within 20 months of initial therapy, while 15 had no evidence of disease with a minimum follow-up of 36 months. Polymerase chain reaction (PCR) was performed on paraffin-block tissue of the hysterectomy specimen cervical tumor and lymph nodes. Oligonucleotide probes for HPV types 6, 11, 16, 18, 31, 33, and 35 were used with consensus primers for uncharacterized HPV types created from an L1 constant region. Control tissues were run with each tumor sample to assure against contamination. HPV type confirmation was performed using diagnostic restriction sites. HPV was detected in all cervical tumors. Recurring tumors were infected with multiple types of HPV in all 6 tumors versus only 5 of 15 nonrecurring tumors being multiply infected (P = 0.023). No tumor had HPV 6 or 11, and the incidence of HPV 16, 31, 33, and 35 was not significantly different for recurrent versus nonrecurrent groups. HPV 18 was found in 5 of 6 recurring cancers versus 1 of 15 nonrecurring tumors (P = 0.0029). PCR typing of the histologically negative nodes that had been obtained at radical hysterectomy was done in all 6 recurring patients and in 6 nonrecurring patients. The recurrent patients had a significantly higher incidence of lymph nodes positive for HPV DNA (71%) than the nonrecurring patients (35%) (P = 0.0047). These observations suggest that HPV 18 cervical cancer patients, those with infections of multiple types, and those with HPV DNA in histologically negative lymph nodes may be at increased risk for recurrence.